GENETICS OF INCIPIENT SPECIATION IN DROSOPHILA MOJAVENSIS: II. HOST PLANTS AND MATING STATUS INFLUENCE CUTICULAR HYDROCARBON QTL EXPRESSION AND G × E INTERACTIONS

We performed a quantitative trait locus (QTL) analysis of epicuticular hydrocarbon variation in 1650 F₂ males from crosses of Baja California and mainland Mexico populations of Drosophila mojavensis cultured on two major host cacti. Principal component (PC) analysis revealed five PCs that accounted for 82% of the total epicuticular hydrocarbon variation. Courtship trials with mainland females were used to characterize hydrocarbon profiles of mated and unmated F₂ males, and logistic regression analysis showed that cactus substrates, two PCs, and a PC by cactus interaction were associated with mating success. Multiple QTLs were detected for each hydrocarbon PC and seven G × E (cactus) interactions were uncovered for the X, second, and fourth chromosomes. Males from the courtship trials and virgins were used, so "exposure to females" was included as a factor in QTL analyses. "Exposed" males expressed significantly different hydrocarbon profiles than virgins for most QTLs, particularly for the two PCs associated with mating success. Ten QTLs showed G × E (exposure) interactions with most resulting from mainland genotypes expressing altered hydrocarbon amounts when exposed to females compared to Baja genotypes. Many cactus × exposure interaction terms detected across QTL and all PCs confirmed that organ pipe-reared males expressed significantly lower hydrocarbon amounts when exposed to females than when reared on agria cactus. Epicuticular hydrocarbon variation in D. mojavensis is therefore a multigenic trait with some epistasis, multiple QTLs exhibited pleiotropy, correlated groups of hydrocarbons and cactus substrates determined courtship success, and males altered their hydrocarbon profiles in response to females.     

Premating isolation is determined by larval rearing substrates in cactophilic Drosophila mojavensis. IX. Host plant and population specific epicuticular hydrocarbon expression influences mate choice and sexual selection

Sexual signals in cactophilic Drosophila mojavensis include cuticular hydrocarbons (CHCs), contact pheromones that mediate female discrimination of males during courtship. CHCs, along with male courtship songs, cause premating isolation between diverged populations, and are influenced by genotype × environment interactions caused by different host cacti. CHC profiles of mated and unmated adult flies from a Baja California and a mainland Mexico population of D. mojavensis reared on two host cacti were assayed to test the hypothesis that male CHCs mediate within‐population female discrimination of males. In multiple choice courtship trials, mated and unmated males differed in CHC profiles, indicating that females prefer males with particular blends of CHCs. Mated and unmated females significantly differed in CHC profiles as well. Adults in the choice trials had CHC profiles that were significantly different from those in pair‐mated adults from no‐choice trials revealing an influence of sexual selection. Females preferred different male CHC blends in each population, but the influence of host cactus on CHC variation was significant only in the mainland population indicating population‐specific plasticity in CHCs. Different groups of CHCs mediated female choice‐based sexual selection in each population suggesting that geographical and ecological divergence has the potential to promote divergence in mate communication systems.     

Premating isolation is determined by larval rearing substrates in cactophilic Drosophila mojavensis. X. Age‐specific dynamics of adult epicuticular hydrocarbon expression in response to different host plants

Analysis of sexual selection and sexual isolation in Drosophila mojavensis and its relatives has revealed a pervasive role of rearing substrates on adult courtship behavior when flies were reared on fermenting cactus in preadult stages. Here, we assessed expression of contact pheromones comprised of epicuticular hydrocarbons (CHCs) from eclosion to 28 days of age in adults from two populations reared on fermenting tissues of two host cacti over the entire life cycle. Flies were never exposed to laboratory food and showed significant reductions in average CHC amounts consistent with CHCs of wild‐caught flies. Overall, total hydrocarbon amounts increased from eclosion to 14–18 days, well past age at sexual maturity, and then declined in older flies. Most flies did not survive past 4 weeks. Baja California and mainland populations showed significantly different age‐specific CHC profiles where Baja adults showed far less age‐specific changes in CHC expression. Adults from populations reared on the host cactus typically used in nature expressed more CHCs than on the alternate host. MANCOVA with age as the covariate for the first six CHC principal components showed extensive differences in CHC composition due to age, population, cactus, sex, and age × population, age × sex, and age × cactus interactions. Thus, understanding variation in CHC composition as adult D. mojavensis age requires information about population and host plant differences, with potential influences on patterns of mate choice, sexual selection, and sexual isolation, and ultimately how these pheromones are expressed in natural populations. Studies of drosophilid aging in the wild are badly needed.     

Evolution of developmental homeostasis inDrosophila mojavensis

Summary Variation in life histories among populations of cactophilicDrosophila mojavensis has been hypothesized to be a by-product of a shift to one of two alternate host plants. When cultured on the ancestral and a secondary host cactus, a Baja population expressed shorter development times and smaller thorax sizes than a mainland population, but viability did not differ. Comparisons with all reciprocal F₁ and F₂ crosses between populations revealed that genetic differences in development time and thorax size were largely additive. Homeostasis in these life history traits was population specific, except for viability. Homeostasis in development time was greater in the Baja population than in the other crosses, suggesting dominance for decreased homeostasis in the mainland population. Underdominance in viability homeostasis of the F₁ hybrids suggested some incompatibility between populations. Homeostasis in thorax size was greater in females than in males and differed among parental populations. Maintenance of heritable differences and genetic variation for homeostasis in these traits suggested a role for cactus-specific differences in environmental uncertainty caused by variation in breeding site duration and abundance in nature.     

Pre-mating isolation is determined by larval rearing substrates in cactophilicDrosophila mojavensis. II. Effects of larval substrates on time to copulation,mate choice and mating propensity

Summary It has been hypothesized that reproductive character displacement has evolved in mainland Sonora, Mexico populations of cactophilicD. mojavensis due to the presence of a sympatric sibling speciesD. arizonae. In laboratory tests using ancestral Baja California populations and derived, sympatric mainland populations, asymmetrical sexual isolation has been observed among populations ofD. mojavensis where mainland females discriminate against Baja males. Effects of different pre-adult rearing environments on adult mating behaviour were assessed by comparing fermenting cactus tissues like those used in nature for breeding with laboratory media because previous studies have employed synthetic growth media for fly growth and development. Significant behavioural isolation was evident in all cases when larvae were reared on laboratory food, but was non-significant when flies were reared on fermenting cactus, except for the cactus used by most mainland populations, consistent with previous studies. Time to copulation of Baja females was greater than mainland females over all substrates, but male time to copulation did not differ between populations. Time to copulation for both sexes was significantly greater when flies were reared on laboratory food with one exception. The degree of behavioural isolation was weakly correlated with time to copulation across food types (Spearman rank correlation = 0.58,p = 0.099). Therefore, use of laboratory media in this and previous studies exaggerated adult pre-mating isolation and time to copulation in comparison to natural breeding substrates. These experiments suggest that a change in host substrates by saprophagous insects (where chemical differences exist between hosts) may have subtle effects on mating behaviour in a manner which promotes low levels of sexual isolation as a by-product of their utilization of a particular substrate during larval development. ForD. mojavensis, these results suggest that over evolutionary time, radiation into a new environment (from Baja to the mainland) allowed utilization of new host plants that may have incidentally promoted the sexual isolation patterns that have been observed within this species.See Etges (1992) for the first paper in this series.     

Post‐zygotic isolation in cactophilic Drosophila: larval viability and adult life‐history traits of D. mojavensis/D. arizonae hybrids

Drosophila mojavensis and Drosophila arizonae are cactophilic flies that have been used extensively in speciation studies. Incomplete premating isolation, evidence of reinforcement, and a lack of recent introgression between these species point to a potentially important role for post‐zygotic isolating barriers in this system. Other than hybrid male sterility, however, post‐zygotic isolation between D. mojavensis and D. arizonae has received little attention. In this study, we examined viability and life‐history traits of D. mojavensis/D. arizonae F₁ hybrids from sympatric crosses. Specifically, we reared hybrids and purebreds on the natural host cacti of each parental species and compared viability, development time, thorax length, and desiccation resistance between hybrids and purebreds. Interestingly, hybrid females from both crosses performed similarly or even better than purebred females. In contrast, hybrid sons of D. arizonae mothers, in addition to being sterile, had shorter average thorax length than males of both parental species, and hybrid males from both crosses had substantially lower desiccation resistance than D. mojavensis males. The probable cost to hybridization for D. mojavensis females resulting from reduced desiccation resistance of hybrid sons may have been an important selective factor in the history of reinforcement for crosses involving these females.     

Premating isolation is determined by larval-rearing substrates in cactophilic Drosophila mojavensis. V. Deep geographic variation in epicuticular hydrocarbons among isolated populations

Adult epicuticular hydrocarbon variation of 14 geographically isolated populations of cactophilic Drosophila mojavensis was assessed to further investigate mechanisms of sexual isolation. Hydrocarbon transfer experiments demonstrated that these compounds are part of the mate recognition system in this species. Sixteen of the 23 epicuticular hydrocarbon components studied differed in amounts between males and females, and 13 differed in quantity between the geographic regions encompassing Baja California and mainland Mexico (Sonora and Sinaloa). Eight hydrocarbon components, seven of which differed in quantity between sexes, showed significant sex-by-region interactions, indicating region-specific sex reversals in hydrocarbon quantities. Such regional variation in epicuticular hydrocarbon profiles suggests that these hydrocarbon differences have also evolved in D. mojavensis since this species invaded mainland Sonora and Sinaloa from Baja California by switching host plants, in addition to a number of key genetic, behavioral, and life-history characters.     

ALTERED MATING BEHAVIOR IN A CARSONIAN POPULATION OF DROSOPHILA SECHELLIA

Mating behavior was studied in two laboratory populations of Drosophila sechellia and their reciprocal hybrids. The ancestral population was reared on a special medium, optimal for this species, while the derived population was reared on a standard Drosophila food, and underwent a bottleneck while adapting to this new medium, in a manner similar to the “founder-flush” process of Carson (1971). A significant tendency towards mating asymmetry was found, with ancestral females mating significantly less frequently with derived males than derived females with ancestral males. Analysis of hybrids suggested an important role for the male's X chromosome or for a maternal effect. No significant differences were found among parental types for their main female cuticular hydrocarbons, the proportion of courtship spent in various behavioral elements, body weight, or wing length. Significant differences were found in the structure of courtship, male locomotor activity, male cuticular hydrocarbon levels, and male courtship song inter-pulse interval (i.p.i.). None of these differences showed an X-linked effect in the reciprocal hybrids. Hypotheses put forward to explain interspecific mating asymmetries are discussed in the light of these results.     

Sensitivity to larval density in populations of Drosophila mojavensis: Influences of host plant variation on components of fitness

Summary Chromosomally polymorphic populations of Drosophila mojavensis from Baja California feed and breed on agria cactus, Stenocereus gummosus; whereas, monomorphic Arizona populations are associated exclusively with organ pipe cactus, S. thurberi. The effects of this host plant shift in expanding the kinds of feeding and breeding sites were assessed by manipulating larval density and recording differences in egg to adult development time and viability, and adult thorax size in both populations on artificially rotted substrates of both cactus species. Older agria rots increased development time but had no effect on viability. Organ pipe rots were qualitatively poorer substrates than agria rots for both monomorphic and polymorphic populations of D. mojavensis, especially at higher larval densities causing longer egg to adult development times, lower viabilities, and smaller thorax sizes than agria.The Baja population expressed shorter development times, higher viabilities, and smaller thorax sizes than the Arizona population on both cactus substrates. No evidence for cactus host race formation was found. The Baja population was less sensitive to increasing larval densities for all fitness characters studied on both cactus substrates indicating greater developmental homeostasis than in the monomorphic Arizona population. These data support the hypothesized central-marginal population structure within this species coincident with the distribution of host plants and lend insight into the process of adaptive divergence at different life history stages caused by host plant shifts.     

GENETICS OF HOST-CACTUS RESPONSE AND LIFE-HISTORY EVOLUTION AMONG ANCESTRAL AND DERIVED POPULATIONS OF CACTOPHILIC DROSOPHILA MOJAVENSIS

The extent of host-specific genetic variation for two life-history traits, egg to adult developmental time and viability, and one morphological trait closely tied to fitness, adult thorax size, was exposed by employing a nested half-sib/full-sib breeding design with Baja and mainland populations of Drosophila mojavensis recently extracted from nature. This study was motivated by the presence of substantial variation in life histories among populations of D. mojavensis that use the fermenting tissues of particular species of columnar cacti for feeding and breeding in the Sonoran Desert. Full-sib progeny from all sire-dam crosses were split into cultures of agria cactus, Stenocereus gummosus, and organ pipe cactus, S. thurberi, to examine patterns of genotype-by-environment interaction for these fitness components. Baja flies expressed shorter egg-to-adult developmental times, higher viabilities, and smaller body sizes than mainland flies consistent with previous studies. Significant sire and dam components of variance were exposed for developmental time and thorax size. Genotype-by-environment interactions were significant at the level of dams for developmental time and nearly significant for viability (P = 0.09). Narrow- and broad-sense heritabilities were influenced by host cactus, sex, and population. No strong pattern of genetic correlation emerged among fitness components suggesting that host-range expansion has not been accompanied by formation of coadapted life histories, yet the ability to estimate genetic correlations and their standard errors was compromised by the unbalanced nature of the data set. Genetic correlations in performance across cacti were slightly positive, evidence for ecological generalism among populations explaining the observed pattern of multiple host cactus use within the species range of D. mojavensis.     

Genetics of incipient speciation in Drosophila mojavensis. I. Male courtship song, mating success, and genotype x environment interactions

Few studies have examined genotype by environment (GxE) effects on premating reproductive isolation and associated behaviors, even though such effects may be common when speciation is driven by adaptation to different environments. In this study, mating success and courtship song differences among diverging populations of Drosophila mojavensis were investigated in a two-environment quantitative trait locus (QTL) analysis. Baja California and mainland Mexico populations of D. mojavensis feed and breed on different host cacti, so these host plants were used to culture F2 males to examine host-specific QTL effects and GxE interactions influencing mating success and courtship songs. Linear selection gradient analysis showed that mainland females mated with males that produced songs with significantly shorter L(long)-IPIs, burst durations, and interburst intervals. Twenty-one microsatellite loci distributed across all five major chromosomes were used to localize effects of mating success, time to copulation, and courtship song components. Male courtship success was influenced by a single detected QTL, the main effect of cactus, and four GxE interactions, whereas time to copulation was influenced by three different QTLs on the fourth chromosome. Multiple-locus restricted maximum likelihood (REML) analysis of courtship song revealed consistent effects linked with the same fourth chromosome markers that influenced time to copulation, a number of GxE interactions, and few possible cases of epistasis. GxE interactions for mate choice and song can maintain genetic variation in populations, but alter outcomes of sexual selection and isolation, so signal evolution and reproductive isolation may be slowed in diverging populations. Understanding the genetics of incipient speciation in D. mojavensis clearly depends on cactus-specific expression of traits associated with courtship behavior and sexual isolation.     

MALE HYBRID STERILITY IN DROSOPHILA: INTERACTIONS BETWEEN AUTOSOMES AND SEX CHROMOSOMES IN CROSSES OF D. MOJAVENSIS AND D. ARIZONENSIS

By backcrossing hybrids from the cross Drosophila mojavensis female × Drosophila arizonensis male to both parental species we show that several interspecific combinations of autosomes with one or the other sex chromosome (X or Y) result in sperm abnormalities. Two of these incompatibilities will cause the same type of nonreciprocal F₁ male sterility that is observed in this pair of species, but the possibility of an additional incompatibility that would have the same result, e.g., an incompatibility between the mojavensis Y and the arizonensis X chromosomes, cannot be excluded. The incompatibility between the arizonensis Y chromosome and the mojavensis fourth chromosome found to occur for all tested populations of mojavensis race B (Vigneault and Zouros, 1986) is shown also to occur for race A of this species. We further show that a dominance relationship exists between heterospecific homologous autosomes in their interactions with the sex chromosomes and that the direction of the dominance depends on whether the sex chromosome is the X or the Y. The present role of these incompatibilities in preventing gene flow between the two species may be minor, but their genetic basis and mode of action may provide useful insights about the genetic events that have played a significant role in earlier stages of speciation.     

A comparison of behavioural change in Drosophila during exposure to thermal stress

In order to understand how adaptive tolerance to stress has evolved, we compared related species and populations of Drosophila for a variety of fitness relevant traits while flies directly experienced the stress. Two main questions were addressed. First, how much variation exists in the frequency of both courtship and mating among D. melanogaster, D. simulans, and D. mojavensis when each are exposed to a range of temperatures? Second, how does variation in these same behaviours compare among four geographically isolated populations of D. mojavensis, a desert species with a well defined ecology? Our hierarchical study demonstrated that mating success under stress can vary as much between related species, such as D. melanogaster and D. simulans, as between the ecologically disparate pair, D. melanogaster and D. mojavensis. Strains of this latter desert species likewise varied in tolerance, with differences approaching the levels observed among species. The consequences of stress on male courtship differed markedly from those on female receptivity to courtship, as mating behaviours among species and among strains of D. mojavensis varied in subtle but significant ways. Finally, a comparison of variation in thermotolerance of F₁ hybrids between the two most extreme D. mojavensis populations confirmed that genetic variation underlying traits such as survival or the ability to fly after heat stress is completely different. © 2004 The Linnean Society of London, Biological Journal of the Linnean Society, 2004, 83 , 197–205.     

SELECTION FOR SEXUAL ISOLATION BETWEEN GEOGRAPHIC FORMS OF DROSOPHILA MOJAVENSIS. I. INTERACTIONS BETWEEN THE SELECTED FORMS

Drosophila mojavensis comprises three geographic forms occurring in the United States, the Baja California peninsula, and mainland Mexico. Peninsular and mainland forms were selected for increased sexual isolation from each other, while controls were maintained with maximum outbreeding. Response to selection was highly asymmetrical in that isolation was very high between selected peninsular males and mainland females, but nonexistent between selected mainland males and peninsular females. The heightened isolation is primarily due to some change in the peninsular males.     

Population specific fitness response of Drosophila subobscura to lead pollution

Abstract Differences in heavy metal tolerance among separate populations of the same species have often been interpreted as local adaptation. Persistence of differences after removing the stressor indicates that mechanisms responsible for the increased tolerance were genetically determined. Drosophila subobscura Collin (Diptera: Drosophilidae) populations were sampled from two localities with different history of heavy metal pollution, and reared for eight generations in the laboratory on a standard medium and on media with different concentrations of lead (Pb). To determine whether flies from different natural populations exposed to the Pb‐contaminated media in the laboratory show population specific variability in fitness components over generations, experimental groups with different concentrations of lead were assayed in three generations (F₂, F₅, and F₈) for fecundity, developmental time, and egg‐to‐adult viability. On the contaminated medium, fecundity was reduced in later generations and viability was increased, irrespective of the environmental origin of populations. For both populations, developmental time showed a tendency of slowing down on media with lead. Faster development was observed in later generations. Preadaptation to contamination, meaning higher fecundity, higher viability, and faster egg to adult development in all studied generations, was found in D. subobscura originating from the locality with a higher level of heavy metal pollution.